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A Split Staphylococcus aureus Cas9 as a Compact Genome-Editing Tool in Plants.
Hidetaka Kaya1, Kazuhiro Ishibashi2, Seiichi Toki1,3,4
1Plant Genome Engineering Research Unit, Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, Kannondai, Tsukuba, Ibaraki, Japan.
Plant & Cell Physiology
|April 4, 2017
Summary
Split-protein Staphylococcus aureus Cas9 (SaCas9) enables precise CRISPR/Cas9 genome editing in plants. This smaller, split SaCas9 tool offers temporal and spatial control for targeted mutagenesis, advancing plant biotechnology.
Area of Science:
- Molecular Biology
- Plant Biotechnology
- Gene Editing
Background:
- Split-protein systems regulate protein activity by requiring reassembly of inactive fragments.
- CRISPR/Cas9 technology is a powerful tool for genome editing.
Purpose of the Study:
- To investigate the feasibility of using split Staphylococcus aureus Cas9 (SaCas9) for targeted mutagenesis in plants.
- To develop a smaller and more controllable CRISPR/Cas9 system for plant genome editing.
Main Methods:
- Expressing split SaCas9 fragments in Nicotiana benthamiana using Agrobacterium and Tomato mosaic virus vectors.
- Assessing genome editing activity through targeted mutagenesis.
Main Results:
- Split SaCas9, specifically the _739N/740C construct, demonstrated significant genome editing activity comparable to full-length SaCas9.
- The use of plant virus vectors for split SaCas9 expression facilitates integration-free genome editing.
Conclusions:
- Split SaCas9 is a viable and effective tool for targeted plant genome editing.
- Split SaCas9 systems offer temporal and spatial regulation of CRISPR/Cas9 activity in plant cells, enabling advanced genome editing applications.